Delivery of siRNA and other therapeutic oligonucleotides is a major technical challenge that has limited their potential for clinical translation.
A liposome is a vesicle composed of one or more lipid bilayers, capable of carrying hydrophilic molecules within an aqueous core or hydrophobic molecules within its lipid bilayer(s). Lipid nanoparticles (LNs) is a general term to described lipid-based particles in the submicron range. They can have structural characteristics of liposomes and/or have alternative non-bilayer types of structures. Drug delivery by LNs via systemic route requires overcoming several physiological barriers. The reticuloendothelial system (RES) can be responsible for clearance of LNs from the circulation. Once escaping the vasculature and reaching the target cell, LNs are typically taken up by endocytosis and must release the drug into the cytoplasm prior to degradation within acidic endosome conditions.
Consideration of zeta potential or surface charge is necessary when preparing LNs. The zeta potential of LNs typically should not be excessively positive or too negative for systemic delivery. LNs with a highly positive charge tend to interact non-specifically with target cells and circulating plasma proteins, and may cause cytotoxicity. Alternatively, LNs with a highly negative charge cannot effectively incorporate nucleic acids, which are also negatively charged, and may trigger rapid RES-mediated clearance, reducing therapeutic efficacy. LNs with a neutral to moderate charge are best suited for in vivo drug and gene delivery.
LNs constitute a promising platform for the delivery of traditional therapeutic compounds and nucleic acid-based therapies. Drugs formulated using LNs can often feature superior pharmacokinetic (PK) properties in vivo, such as increased blood circulation time and increased accumulation at the site of solid tumors due to enhanced permeability and retention (EPR) effect. Moreover, LNs may be surface-coated with polyethylene glycol to reduce opsonization of LNs by serum proteins and the resulting RES-mediated uptake. LNs can also be coated with cell-specific ligands to provide targeted drug delivery.
Much interest has arisen over nucleic acid-based therapies over the past few decades. Nucleic acid-based therapies work on the premise of introducing nucleic acids (NAs) to promote or inhibit gene expression. As mutations in genes and changes in miRNA profile are believed to be the underlying cause of cancer and other diseases, nucleic acid-based agents potentially can directly act upon the underlying etiology, maximizing therapeutic potential. A few examples of nucleic acid-based therapies include plasmid DNA (pDNA), small interfering RNA (siRNA), small hairpin RNA (shRNA), microRNA (miR) mimic (or mimetic), anti-miR/antagomiR/miR inhibitor, and antisense oligonucleotide (ASO), each of which is encompassed by the term nucleic acid as used in the present disclosure. The clinical translation of nucleic acid-based therapies faces several obstacles in its implementation. Transporting nucleic acids to their intracellular target is particularly challenging as nucleic acids are relatively unstable and are subject to degradation by serum and cellular nucleases. Further, the high negative charges of nucleic acids make it impossible for transport across the cell membrane, limiting utility. Viral vectors have been developed to address this issue, but most have failed due to activation of immunological responses in vivo and induction of undesired mutations in the host genome. Non-viral vectors have also been investigated extensively, but few have yielded successful clinical outcomes and further improvements are needed.
Traditionally, cationic LNs have been utilized as non-viral vectors for gene delivery. In some instances, cationic lipids are replaced with, or used in combination with, anionic lipids. The positive charge of cationic LNs facilitates an electrostatic interaction with negatively charged nucleic acids. Anionic lipids can be combined with cationic lipids or with a cationic polymer, which will in turn mediate interaction with the nucleic acids. These may be prepared by various techniques known in the art such as ethanol dilution, freeze-thaw, diafiltration, and thin film hydration. In addition to cationic components, LNs are typically composed of helper lipids, including bilayer-forming phospholipid components such as phosphatidylcholines, as well as cholesterol. Helper lipids such as dioleoylphosphatidylethanolamines (DOPE) do not favor bilayer phase and instead aid in disrupting the lipid bilayer at the target site to release the therapeutic agent. Stabilizing components such as D-alpha tocopheryl polyethylene glycol 1000 succinate (TPGS), which is a PEGylating agent, or mPEG-DSPE may be added to stabilize the formulation and protect the LN from RES-mediated uptake.
The development of efficient delivery vehicles is a key to clinical translation of oligonucleotide (ON) therapeutics. Ideally, a lipid nanoparticle formulation should be able to (1) protect the drug from enzymatic degradation; (2) traverse the capillary endothelium; (3) specifically reach the target cell type without causing immunogenicity or off-target cytotoxicity; (4) promote endocytosis and endosomal release; and (5) form a stable formulation with colloidal stability and long shelf-life.